2. Introduction:
•A blood clot (thrombus) developed in the
circulatory system can cause vascular blockage
leading to serious consequences including
death.
• A healthy hemostatic system suppresses the
development of blood clots in normal
circulation, but reacts extensively in the event
of vascular injury to prevent blood loss.
3. Outcomes of a failed homeostasis include
stroke, pulmonary embolism, deep vain
thrombosis and acute myocardial infraction.
4. STREPTOCOCCAL FIBRINOLYSIS
These enzyme activates a fibrinolytic
enzyme in human serum, which splits fibrin
into smaller fragments, thus it causes rapid
dissolution of blood clots and fibrinous
exudates. Therefore, streptokinase acts
indirectly upon a substrate of fibrin or
fibrinogen.
5. SOURCE
This enzyme is produced by certain bacteria. The
most frequently employed for manufacture are:
haemolytic streptococci, particularly those of the
lancefied groups A, human C and G.
Steptodornase is a related enzyme, which act
directly upon a substrate of
deoxyribonucleoproteins (DNA).
6. These are main constituents of nuclei. They
also present upto 30-70% in purulent
exudates. Steptodornase splits them into free
purine bases and pyrimidine nucleosides
thereby decrese the viscosity of purulent
exudates.
7. MANUFACTURING PROCESS
It is produced by fermentation process ‘
which involves the following steps:-
1. Preparation of medium
2. Fermentation
3. Purification of the product.
8. PREPARATION OF MEDIUM
Ingredients: Casein digest solution: It is prepared by
dissolving casein in water in specified proportion. It is
heated to 100 C and maintained the same
temperature, till the solution is clear. The resultant
solution is rapidly cooled to 15 C and filtered through
a coarse filter paper. Toluene in small quantity is
added for the purpose of preservation. It is stored for
four days at 20 C and filtered to remove insoluble
material.
9. 2. Dextrose (a carbohydrate source)
3. Amino acids
a)Cysteine in 10%HCl
b)Glycine
c)Tryptophan.
4. Vitamins
a)Thiamine hydrocloride
b)Riboflavin
c)Nicotinic acid
d)Pyridoxine
e)Calcium pentothenate
a)
11. FERMENTATION
Sterilized medium is inoculated with seed
inoculation of bacterium; S. haemolyticus
having a bacterial count of 20 billions/ml.
fermentation is carried out in a tank for 14hrs
at 37C. During this period no pH adjustment,
aeration or modification are made.
12. Later, dextrose 50% is added and pH is
adjusted to 6 at 15 min. interval with 5N
sodium hydroxide. After each adjustment of
pH, 50% dextrose is added. Fermentation is
continued till bacterial count ceases to
increase(about 3 hrs).At this stage
fermentation medium contains appx. 1000
units/ml
13. PURIFICATION
Crude streptokinase is first dialysed against
phosphate buffer then it is applied on
modified cellulosic columns and eluted with
phosphate buffer with increasing pH and
molarity(increasing pH 5.8 to 8.5 and
molarity 0.005to 0.1 M).
14. pH and molarity are the important factors in
purification.
At 0.1 M: streptokinase is eluted
> 0.1 M: improper adsorption and separation pH
> 8.5:adsorption capacity of cellulose decreases
< 5.8:streptokinase is precipitated
At pH 8 and molarity 0.75, impurities are eluted.
15. PURIFICATION BY DEAE
CELLULOSE
Crude streptokinase, phosphate buffer 0.2 M and
DEAE cellulose in the proportion of 3:2:1 are stirred
for 1 hour and filtered. Cake is washed with
phosphate buffer 0.025 m by stirring for 30 mins. It
is again filtered and cake is suspended in 0.1 m
phosphate buffer by stirring for 1 hour. Filterate is
collected which contains pure streptokinase.
16. USES
Treatment of thromboembolic disorders for
the lysis of pulmonary emboli, arterial
thrombus, deep vein thrombus and acute
coronary artery thrombosis.